Detection systems and methods

ABSTRACT

The application describes apparatus and methods using ultrasound transmitted through a chromatography column bed to determine the status of materials in the bed space, particularly during chromatography runs. Detected conditions include the profile of a band of eluting material—to see whether it is satisfactory—the arrival of a band of material at the column output, the fitness for use of the ed packing, the passage of different mobile phases, e.g. salt fronts, and other conditions. Aspects claimed include control processors and corresponding programmed products.

REFERENCE TO RELATED APPLICATIONS

This application is a 371 of PCT/GB02/0533 filed Nov. 27, 2002.

FIELD OF THE INVENTION

This invention relates to detection systems and methods used in thefield of chromatography, and to apparatus and methods for chromatographyincorporating such detection systems and methods. Chromatography is theseparation—often for purposes of purification—of components in a mixtureby passing them with an eluent liquid through a bed of particles.

BACKGROUND

It is normal for various detectors to be present in the piping systemsassociated with a chromatography column, either immediately adjacent oron the column, or in associated machinery such as packing skids whichmay from time to time be connected to the column. For example it isnormal for the outflow from the column to include a pH meter and/or a UVmonitor, the latter detecting the presence of chemical species in theoutflow which have UV-absorptive chromophores. Normally a pressure gaugeis connected to the column interior or pumping system.

In the prior-published art, U.S. Pat. No. 4,324,131 describes using aultrasonic detector to detect solutes in the effluent liquid from achromatography column; only schematic details are given but essentiallyultrasound is passed through a cell through which the effluent passesand a second, control cell containing pure eluent; differences in thetransmitted ultrasound amplitude indicate dissolved substances.

Our earlier but not prior-published WO 02/10739, the contents of whichare hereby incorporated by reference, describes a different mode of useof ultrasound in relation to chromatography columns and processes. It isdisclosed that vibratory transmissions, especially ultrasound, can passright across the packed column interior and indicate the situationwithin. A particular aspect is showing the presence of absence of packedbed material (medium); various methods and apparatus are disclosed forusing ultrasound arrangements to monitor and control the packing of acolumn. It was also found that the ultrasound transmissions can detectthe presence of additional substances in the column, in particular bandsof eluted components passing through, and bound impurities or othercomponents remaining in a band on the media so as to affect theultrasound transmission.

THE INVENTION

The present application proposes further developments and refinements ofthe proposals disclosed in WO 02/10739, with particular but notexclusive reference to the use of mechanical vibratory transmissions andespecially ultrasound to detect adventitious material in the column bed,particularly component(s) being separated or purified but also materialwhich may be temporarily or permanently fixed—bound or physicallyretained—on the medium particles in the column. Also, methods in whichthe transmissions are used to detect in-column phenomena associated withprocess runs but not necessarily involving eluted components as such.

Some aspects of the invention are set out in the claims.

In one novel aspect we have found that, as a given substance elutesthrough the column across a signal transmission path, the resultingprofile of modification (attenuation or speed change) of the mechanicalvibratory transmission e.g. ultrasound is to some extent distinctive,i.e. can be used to distinguish a band of a given component from a bandof at least one other different component that might be present with it,or from an improperly-eluting band of the same component.

The exact reasons for the form of these distinctive transmissionmodification profiles are of course complex. They relate to details ofthe interactions between the eluting component, the particulate bedmedium and the eluent. They may reflect the presence of near-identicalsubstances e.g. isomers and variants eluting together through the bed atslightly different rates. Whatever the reasons, we have found that inpractice different substances e.g. different proteins can givecharacteristic traces.

It is possible that the temperature of the bed varies slightly onabsorption and desorption of the eluting substance on and off the mediaparticles. This has the potential to modify the transmission because thepresent techniques allow changes in the column to influence thetransmission over a very long path length; twice the transversedimension of the column which for many industrial-scale columns willgive a path length of a meter or more. However this is one theory and weare not bound to it.

We do note that, where the present technique is used to detect bandprofiles which are also UV-absorptive on leaving the column, there aresimilarities in the band profiles between the two techniques and this ismore readily understood as corresponding to the spreading of peaks intodistinct sub-peaks e.g. because of isomeric forms eluting at slightlydifferent rates. However the present ultrasound transmissions(henceforth the term “ultrasound” used in the present statements of theinvention refers generically to acoustic or mechanical vibratorytransmissions unless the context requires otherwise) can detect thecomponent when it is inside the column, whereas UV requires atransparent path outside the column. Thus, the detection monitoring isavailable throughout the process including important on-column stages,also for substances which do not absorb UV light.

Also, the purity of a sample may be calculated from the ratio of thesizes of peaks in its ultrasound profile. This is a known technique inUV spectroscopy, but has not previously been used with ultrasound. Acontrol processor may be programmed accordingly, or the data interpretedvisually by an operator.

Preferably the detection technique monitors the amplitude/attenuation ofthe transmitted ultrasound rather than speed changes, because the latterare found rather sensitive to temperature which may be disadvantageousin many cases.

In one general apparatus aspect, the present invention relates to theprovision and use of a control processor in relation to a chromatographyapparatus comprising a chromatography column having a housing wall withside wall and end portions defining an internal bed space for containinga particulate chromatography medium, and having at least one transmitterfor transmitting a vibratory mechanical signal such as ultrasoundthrough the bed space, and at least one detector to detect thetransmitted signal(s).

In a first aspect the control processor is programmed to compare atransmission profile observed in the course of a chromatography processwith a model transmission profile for a component of interest, anddetermine future processing conditions or information provided to theuser on the basis of the comparison. It may additionally cause an imageor trace of the profile to be displayed (this is disclosed in ourearlier application). In particular, the control processor—which may bea programmable logic controller (plc) or a pc—may initiate a warning ifthe comparison indicates a mismatch between an actual detected profileand a predetermined model profile for a given component. The revealingof significant differences, for example serious tailing or spreading ofthe profile, or much smaller than predicted area under the profile peakindicating lack of product, can be used as an indication of a faultyprocess perhaps long before the substance reaches the column outlet. Thefault might arise because of a fault in the bed, an improperly loadedsample, or some fault in the preparation of the sample. This early checkenables a run to be terminated early if things are not going properly,potentially avoiding significant wasted time and cost.

The control processor may additionally or alternatively be programmed toinitiate the operation of one or more system valves acting on effluentfrom the column, e.g. switching output from the column from a wasteoutput line to a sample collection output line, in dependence on theimpending arrival at the column outlet, predicted by the detectedultrasound transmission on the column, of a band of an eluted targetcomponent. By tracking progress of the band before it arrives at theoutlet, more accurate timing is achievable and “tighter cuts” than byusing UV alone, downstream of the outlet. This process may or may notinvolve the comparison of the profile with a model profile as proposedabove. Additionally or alternatively, the control processor may simplyinitiate a warning signal in dependence on the impending arrival of acomponent at the column outlet detected as described.

Where appropriate, stored data representing model component profilesunder the appropriate conditions may be provided on removable datacarriers e.g. magnetic cards or disks which can be read by the controlprocessor or by a reader associated with it.

In a preferred arrangement, the chromatography apparatus provides aseries of transmission paths distributed axially along thechromatography column, e.g. by means of an array of transmitters and/orsensors, and data are gathered by periodically taking a set of readingsover the set of paths in a short period, e.g. as a “scan” up or down thewhole or a selected part of the column. Preferably at least 5, morepreferably at least 10 or at least 15 sensors are used.

The periodicity of the multi-path scan is not critical and may be suitedto the process in hand. This technique is appropriate for all on-columnapplications of ultrasound, incidentally, and not restricted to thosedetecting eluting bands of product.

Ultrasound detection may be used with a wide variety of elutingmolecules, from simple small molecules—which may include solventmolecules—through larger molecules to linear and globular molecules suchas proteins and globulins, for example immunoglobulins which may be forexample monoclonal antibodies. These are high-value products inbiochemical processing and we find that our ultrasound techniques detectthem well.

Interestingly, the ultrasound methodology enables detection of someevents to which conventional spectroscopy e.g. UV would be blind, evenif it could see into the column. In particular, the ultrasound issensitive to changes of pressure or compression in the column. Thus, itcan be seen from an ultrasound trace of a run when the column isswitched offline. It can also be seen where the eluent e.g. elutingbuffer is changed, for example to wash from the column components whichpreviously were bound. Without being committed to the theory, this maybe because different eluent liquids have different viscosities, so thatthey compress the bed to varying degrees as they are pumped through. Theultrasound record of a run showing such events is a valuable diagnostictool.

Another valuable tool is to indicate the base line change relative tothe “clean” medium before the first run. Successive runs progressivelyaccumulate bound material on the media particles which may not all beable to be washed off. Acceptable threshold levels of such boundmaterial may be determined and an initial ultrasound check or scan ofthe bed used to check whether the base line binding is acceptable. Ourearlier application disclosed this for a band of impurity at the inputside of the column. It is disclosed here in relation to the column as awhole, or to parts of the bed spaced away from the input.

The ultrasound technique may provide new and useful ways of testing thepurification procedures used as well as the purity of the productsthemselves. Ultrasound can be used to check the purity and concentrationof the sample as outlined above, or it can be used to test the‘goodness’ of the pack without the need (as is conventional) to pass atest chemical (e.g. salt or acetone) through the column. The ultrasoundmay be able to look at a pack directly to assess how well it is packed.This is desirable given the cost of such media; the ability to produce atested, yet unused, column may be a commercial advantage.

A further use benefit in is in being able to see while still on thecolumn when (where, axially) given components become effectivelyresolved. In pilot stages this enables a rapid determination as to thelength of bed required for resolving given components under givenconditions.

A further aspect relates to a sensor array. Because of the usefulness ofthe technique and its applicability to existing columns, a preferredaspect of the invention is an array of ultrasound sensors (transmittersand/or detectors) configured to be secured in an operating positionagainst the side wall of a chromatography column, by means of fasteningmeans which are preferably integrally joined to the array, and which isreleasable so that the array can be transferred from one column toanother. The array of ultrasound sensors preferably has an electricalconnector for the sensors which can readily be disconnected e.g.unplugged, e.g. as a single cable, for connection to different controlprocessors. Or, it may be integrally (permanently) joined to a givencontrol processor. The ultrasound array is preferably a single array oftransceivers, although complementary arrays of transmitters anddetectors may be used instead. The array device may have a concave frontface shaped for conformity with a column side wall. In one preferredembodiment the transmitter-mounting element has a front recess which isclosed off by the column wall to form a fluid chamber to hold a contactfluid; this has been found to facilitate calibrating and zeroing ofmultiple transmitters, which otherwise are very difficult to bring intoprecisely matched contact with the solid column wall.

We prefer to provide control units for the column packing operation(described in our earlier application) and the column running operation(such as described herein) separately, because it is not normallypreferred to keep pumping skids etc. near the column during a run. So,in a preferred aspect a control unit having the run processing featuresdescribed herein or in the earlier application does not include anypacking pump control programming, or the control apparatus does notinclude any packing pump or pump connections, or is not adapted forpumping packing material. Or, any combination of these absences.

A further aspect of the invention is a computer program product orprogrammable logic controller, embodying software code which, when run,carries out any one or more of the control processing functionsdescribed herein.

Finally, we have noted that the ultrasound methods and apparatusproposed herein and in our earlier application are useful not only onclose-packed but also in loose-packed beds, particularly beds which areexpanded in use in so called expanded-bed chromatography processes.Because the effect on the ultrasound transmission is heavily dependenton the closeness of packing/compression of particles in the bed space,the system may be used in an expanded bed process to indicate the degreeof expansion of the particles within. This is a valuable facilitybecause for a given process there are normally optimal degrees ofexpansion of the bed.

BRIEF DESCRIPTION OF THE DRAWINGS

Experiments, methods and apparatuses explaining and illustrating theinvention and embodiments of it now follow, with reference to theaccompanying drawings in which:

FIG. 1 shows a chromatography column with a single ultrasoundtransceiver installed;

FIG. 2 shows the typical nature of a received ultrasound signal;

FIG. 3 resembles FIG. 2 but showing the detection of a bed fault;

FIG. 4 is an ultrasound trace over time showing the detection ofdifferent proteins eluting through a column;

FIG. 5 is a trace for two passes of different amounts of the sameprotein (egg albumin) showing characteristic features;

FIGS. 6 to 10 show readings during elution of a monoclonal IgG on theFIG. 1 column, partly supplemented with UV readings;

FIGS. 11 and 12 illustrate schematically a multi-transceiver array on acolumn, with a control processor, alarm and display used in achromatography run;

FIG. 13 shows the outputs from a sixteen-transceiver array as aninjection of acetone is passed through a column;

FIG. 14 is a perspective partial view illustrating a detachablemulti-sensor array, and FIG. 15 is a fragmentary view of part of amulti-sensor array adapted to form a fluid chamber.

FIG. 1 shows a pilot set-up for transmitting ultrasound across achromatography column; a commercially available piezoelectrictransceiver is clamped against the column wall which here is atransparent polymeric wall enabling visual cross-reference ofconditions. This apparatus is used in the IgG experiment later on.

FIGS. 2 and 3 illustrate the behaviour of ultrasound transmissionsacross the bed space. Essentially, ultrasound is reflected at interfacesbetween materials of differing densities. There is an immediatereflection (peak P) from the wall immediately adjacent the transducer.Then, after a time corresponding to the time for the ultrasound to crossthe bed space and back again, there are reflected signals Q, R bouncingrespectively from the inner and outer surfaces of the opposite wall. Thecontrol software is programmed to work with the opposite inner wallsignal Q.

FIG. 3 shows how some intervening discontinuity can be “seen” by theultrasound: a flaw created in the centre of the bed shows up as anintervening peak S. This peak was seen to disappear as buffer was passedover time, gradually resolving the flaw.

Experiment 1

In experimental work, we prepared a 400 mm diameter column 200 mm long,loaded with Sepharose 4FF gel. In this work we monitored the amplitudeattenuation of the ultrasound transmission along a single transmissionpath halfway down the column.

The mobile phase was water. With the column stable, one liter of 1% w/valbumin in water was applied to the column. The resulting ultrasoundtrace (x axis time, y axis transmitted signal amplitude) is shown in theupper part of FIG. 4. It shows a particular peak profile for the passageof the albumin band across the detected ultrasound transmission path.Repeated injections of albumin gave profiles of the same shape.

Then, one liter of 1% casein in water was applied to the column andpassed at the same rate. The resulting ultrasound trace is seen in thelower part of FIG. 4. There has been some baseline shift relative to thealbumin run, but the peak profile for the casein was reproducible.Furthermore it was clearly distinct from the albumin peak profile.Without wishing to be bound by theory, it may be that various slightlydifferent forms of casein in the sample eluted at slightly differentrates, leading to the characteristic profile.

The person skilled in the design of electronic process control apparatuswill readily appreciate that these profiles can be stored inmachine-readable electronic or magnetic form, e.g. on a card or disk, orin a PC or PLC (programmable logic controller) and that a controlprocessor can readily be programmed to compare profiles measured in realtime against the stored profiles by means of appropriate “goodness offit” mathematical tests. According to criteria which can be determinedempirically, with reference to chromatography runs using known materialsand with successful results, these comparisons can then be used for theuseful purposes described above.

Experiment 2

In a further experiment, two injections of albumin (first 1 and then 0.5of the same 10 g/l solution) were made successively into a columnrunning under the same conditions. FIG. 5 shows the resulting trace.This shows firstly the reproduction of the characteristic trace profilefor albumin. Secondly it shows a shifting of the transmission baselinefollowing each successive injection. This appears to be attributable tothe progressive retention of a certain amount of sample on the bedmedium, and therefore shows that in general the present system may alsodetect ongoing levels of “fouling” of the column. Thus, practicalthreshold levels of fouling may be correlated by testing with “baseline”transmission levels, and the system then programmed on the basis of thecorrelation to give an indication of the level of fouling and/or awarning as to when a threshold level is reached or approached.

Experiment 3

In this experiment, a mammalian cell culture in a 500-liter fermenterwas used to biosynthesise a monoclonal antibody Immunoglobulin G (IgG).The cell culture supernatant was filtered and two runs totalling 85 gIgG in 200 liters of this filtered liquid was purified on a 180 mmdiameter by 150 mm bed height, 4FF-Sepharose Protein A chromatographycolumn. An ultrasound monitor was placed at the base of the column andthe changes in ultrasound signal were recorded during 2 process runs. Ablank run was also carried out to reveal the effect of buffer changes onultrasound signals. The blank was subtracted from the test to confirmwhich peak was the IgG peak, similar to a ‘spike’ experiment.

The experiment was carried out using a mammalian cell culture; Lonza'sstrain 6A1(100)3. It was fermented in a 500 liter stirred tankfermenter. At the end of the fermentation the cells die and releasetheir IgG; the IgG being designated B72.3.

After fermentation the fermentation liquor was disc stack centrifugedgiving the primary recovery liquid. This was then microfiltered througha ‘Cuno’ depth filter then sterile filtered through a Millipore PVDF 0.2um filter to yield the 200 liter yellow liquid for chromatography. Thisliquid contained 0.425 g IgG/liter or in the total 200 liters, 85 g ofIgG.

A 180 mm diameter Moduline II glass column was packed to 150 mm bedheight with rmp Protein A Sepharose 4 Fast Flow, this being 3.82 liter.The recombinant (rmp) Protein A (from E. coli) from Amersham Biosciences(“APB”) is designed for therapeutic applications that require extremelypure eluates. The rmp Protein A has an r.m.m. of 44.6 kd and containsfive antigen binding domains. On average a single rmp Protein A moleculecan bind two antibody molecules. The binding capacity of the media at200 cm/hr is about 15 mg IgG/ml of media (see published APB datafile18-1141-34 AA, 2000-01).

An Amersham BioProcess System (no. 1357) was used to dilute and supplybuffers and samples to the column and to collect the fractions. Thischromatograph had eight buffer inlet ports (switchable) two pumps, 8pump heads, pre-column conductivity and pressure, bubble trap and filter(by-passable), valves to change column flow direction and by-pass; postcolumn conductivity, pH, flow and UV spectrometer. The data from thesensors was logged to a PC.

The Tables below itemise the chromatography steps performed in the twoexperimental runs.

TABLE 1 showing Run 1: Chromatography Conditions used to Separate IgGfrom Clarified Cell Supernatant Mobile Phase Column Step (150 cm/hr; 640ml/min) Purpose Volumes 1 6 M Guanidine HCl To clean the  8 (aq., pH ca.4.6) column prior to process 2 0.05 M Glycine-Glycinate Equilibrium 10(pH 8.4) buffer 3 Clarified process sample Load 16 (62 l) 4 0.05 MGlycine-Glycinate Wash 10 (pH 8.4) 5 0.1 M Glycine (HCl) Elute IgG  3(pH 3.5) 6 0.1 M Citric Acid Regeneration  2 (pH 2.1) (strip)

TABLE 2 showing Run 2: Chromatography Conditions used to Separate IgGfrom Clarified Cell Supernatant Mobile Phase (150 cm/hr; 640 ml/min +Column Step some variations) Purpose Volumes 1 6 M Guanidine HCl Toclean the  8 (aq., pH ca. 4.6) column prior to process 2 0.05 MGlycine-Glycinate Equilibrium 10 (pH 8.4) buffer 3 Clarified processsample Load 36 (138 l) 4 0.05 M Glycine-Glycinate Wash 10 (pH 8.4) 5 0.1M Glycine (HCl) Elute IgG  3 (pH 3.5) 6 0.1 M Citric Acid Regeneration 2 (pH 2.1) (strip)

A single 1 MHz transducer was placed at the base of the column, as shownin FIG. 1. In this position the ultrasound signal would ‘sense’conditions at the portion of the column bed near the outlet.

FIG. 1 shows the position of the ultrasound transducer clamped at thebottom-right of the column. This picture also shows the process liquidadded at the top of the bed. Note that the bed was compressed as thesample was applied due to its higher viscosity. This change in bedheight caused a change in the ultrasound signal. Such changes inultrasound signal occurred also when high salt plugs were applied to thetop of the column in other experiments on this column.

FIG. 6 shows the ultrasound readings for the entire run. During loadingthe signal strength reduced over time. This is due to the graduallyincreasing loading of the protein A on the medium, making for moreattenuative conditions. The UV (top thin trace) is constant because itis monitoring a dynamic flow situation, where a more or less constantamount of material is immobilised from the stream whilst flowing throughthe column. This makes the UV essentially blind to what is happeninginside the column and the first indication of the column being fullyloaded is the UV going off scale. With the ultrasound we have a view ofthe degree and extent of loading throughout the column, if >1transceiver is used, rather than a column-wide average given by themedia manufacturers. This will allow more efficient loading of a column.

After loading, the column is washed to remove as much un-bound impurityas possible; this can be seen coming off as a tight band A on FIG. 6.Both the UV and ultrasound could see this. On closer inspection theultrasound peak is seen before the UV peak. Because of the tightness ofthis peak versus the dimensions of the transducer it is expected that asmaller transducer would resolve the peak much more clearly. Theultrasound signal levels off after the initial large peak indicatingthat bound species are left on the column, requiring a further pH changeto remove them.

FIG. 7 shows the trace in more detail from the end of the loading phase.Point B shows the column bed relaxing as the column is taken off-line,when the column is put back on-line when the system is full of elutionbuffer the bed can be seen to compress again at C. This is where theblank trace P is overlaid i.e. a trace from a corresponding run where abuffer change was performed on an unloaded bed, so that the antibodytrace could be conclusively separated from the background. The antibodypeak E in the true run can be clearly seen as separate and coming afterthe buffer exchange peak D. Between the peaks there is evidence thatthere is a strong interface between the antibody band and the precedingbuffer exchange interface. The UV antibody peak E′ comes offsignificantly after the ultrasound peak, and they both have much thesame morphology.

The shoulders X, Y that are present on the front flank of the blanktrace and the front flank of the antibody trace appear to be miniaturebuffer exchange interfaces. After the peaks have come off, the tracereturns to the stable level of the elution buffer, which is slightlyless than that of the equilibration buffer.

FIGS. 8 and 9 are supplementary data for the same run, explained in thelegends on the figures.

FIG. 10 shows a second run that loaded. the column much more highly.Again the UV and ultrasound peaks are very similar (i.e. double peaks)with the ultrasound peak viewed before the UV.

These experiments show that the ultrasound equipment can retrieve bothqualitative and quantitative data from within a column packed withmedia, under usage conditions with commercially important molecules.

APPARATUS PROPOSALS

FIG. 11 shows schematically a chromatography process set-up, with anarray of multiple ultrasound transceivers 20 installed right up the sidewall 7 of the column 5. Output from the transceivers 20 is fed torespective inputs of a process control unit 4, preferably a programmedlogic controller chip, which is connected in turn to drive a display 3(indicating pictorially the presence of peaks etc. in the columninterior according to axial position h), an alarm “A” which is soundedwhen comparison of prevailing actual conditions with pre-loaded modelconditions reveals a mismatch above a permissible level, and/or when thesensor indicates (see FIG. 12) that a band 6 of a target substance isnearing the outlet at the bottom of the column, and a connection to theoperating driver for a switching valve 8 which switches output from thecolumn between a collecting line C and a waste line W.

FIG. 13 shows output actually obtained with a multi-sensor array. Thiswas a 400 mm diameter column, 310 mm bed height, the bed Sepharose 6fast flow, the mobile phase water. Sixteen piezoelectric ultrasoundtransducers supplied by Agfa were secured against the outside wall ofthe column in a vertical array. The Y-axis is gain (used to restore thedetected signal to 80% of the original strength, and thereforeindicating the degree of attenuation by passing through the column)while the X-axis is time. The mobile phase was water, and the detectedcomponent was a 1.5 liter injection of 1% acetone. The data indicate thepassage of the acetone peak past the respective transceivers, thedistinctive shape of the acetone peak and the gradual spreading as itmoves down the column.

FIG. 14 shows schematically a portable transceiver array 2, consistingof a plastics unit 23 with sixteen individual transceivers 201 extendingvertically—in this embodiment they are staggered to accommodate theirdimensions—and having securing straps 26 to hold the concave front face22 of the array housing 23 firmly against the outside wall of acorresponding column. A communication cable 25 carries the power andsignals to and from the transducers 201.

FIG. 15 shows a refinement, with the front of the transducer housing 23being recessed to provide an internal cavity 35. The contoured frontedge 31 of the housing has a seal gasket 32 so that, with the housingclamped against the column side wall 7, an enclosed chamber is formed.This can be filled with water or other suitable liquid 28 through a topfill opening 29. This is found to be useful in enabling the zeroing ofthe multiple transducers 201, avoiding difficulties which otherwisearise in getting precise matching of the solid-solid interfaces betweentransducer and column wall.

1. A chromatography apparatus comprising a chromatography column havinga housing wall with side wall and end portions defining an internal bedspace for containing a particulate chromatography medium, and having atleast one transmitter for transmitting vibratory mechanical signalsthrough the bed space, at least one detector for detecting suchtransmitted signals, and a control processor operatively connected tothe transmitter/detector arrangement to detect and input the signalstransmitted through the bed space, the control processor having any oneor more of the following features: (i) being programmed to compare atransmitted signal profile, arising from passage of a band of elutedmaterial through a packed bed in said bed space, with a model signalprofile for such material stored in or accessible to the controlprocessor, and to initiate a warning signal if the comparison indicatesa mismatch between the actual profile and the model profile; (ii) beingprogrammed to initiate the operation of system valves to switch liquidoutput from the chromatography column from a waste output line to asample collector line, in dependence on the impending arrival, indicatedby detected transmissions adjacent the downstream end of the column, ofa band of an eluted component at the column outlet; (iii) beingprogrammed to initiate a warning signal in dependence on the impendingarrival, detected by a said transmission adjacent the downstream end ofthe column, of a band of eluted component at the column outlet; (iv)being programmed to indicate the degree of expansion of an expanded bedinside the bed space, relative to a packed condition.
 2. Thechromatography apparatus according to claim 1 in which an array ofplural ultrasound transmitters is provided distributed axially along thechromatography column.
 3. The chromatography apparatus according toclaim 2 in which the ultrasound transmitters are ultrasoundtransceivers.
 4. The chromatography apparatus according to claim 1 inwhich the control unit is programmed to compare a base line level ofsignal transmissibility with an acceptable threshold level indicatingadequate purity of the particulate medium in the column.
 5. Thechromatography apparatus according to claim 2 in which the ultrasoundtransmitters are detachably secured to the column wall.
 6. Thechromatography apparatus according to claim 1 in which the control unitcomprises a programmable logic controller or personal computer.
 7. Thechromatography apparatus according to claim 1 in which the detectedproperty of the signal is the relative amplitude of the signal.
 8. Thechromatography apparatus according to claim 3 in which the ultrasoundtransmitters are detachably secured to the column wall.
 9. Thechromatography apparatus of claim 1, wherein the control processor hastwo or more of features (i) through (iv).
 10. The chromatographyapparatus of claim 1, wherein the control processor has three or more offeatures (i) through (iv).
 11. The chromatography apparatus of claim 1,wherein the control processor further comprises the feature of beingprogrammed to control the sending of said signals along a plurality oftransmission paths distributed axially along the chromatography column,by initiating periodic scans each being a set of transmissions alongeach of said plural transmission paths.